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一种基于香豆素的靶向基质金属蛋白酶-2的荧光共振能量转移探针,用于宫颈癌检测。

A coumarin-based fluorescence resonance energy transfer probe targeting matrix metalloproteinase-2 for the detection of cervical cancer.

作者信息

He Guangjie, Yang Lu, Qian Xinlai, Li Jing, Yuan Zhiqing, Li Changzheng

机构信息

Department of Forensic Medicine, Xinxiang Medical University, Xinxiang, Henan 453003, P.R. China.

School of Basic Medical Science, Xinxiang Medical University, Xinxiang, Henan 453003, P.R. China.

出版信息

Int J Mol Med. 2017 Jun;39(6):1571-1579. doi: 10.3892/ijmm.2017.2974. Epub 2017 May 3.

DOI:10.3892/ijmm.2017.2974
PMID:28487974
Abstract

Cervical cancer is one of the most common gynecological malignancies worldwide. At present, the methods used for cervical cancer screening have many disadvantages. Matrix metalloproteinase-2 (MMP-2) is low or absent in normal cervical epithelial cells while overexpressed in cervical intraepithelial neoplasia and cervical cancer, which provides the possibility for the detection of cervical cancer based on MMP-2. The development of a strategy with high sensitivity and specificity for cervical cancer detection would be expected to improve the cure rate. Connecting a fluorophore [7-(diethylamino)-2-oxo-2H-chromene-3-succinimidyl ester] with a quencher {4-[4-(dimethylamino)phenylazo]benzoic acid N-succinimidyl ester} via an MMP-2 substrate peptide GPLGVRGKGG, a new coumarin-based fluorescence resonance energy transfer probe targeting MMP-2 was prepared to examine cervical cancer by cell imaging. The cervical cancer cell lines showed green fluorescence with intensities in the order of CaSki > SiHa > C33A > HeLa, which indicated that the cervical cancer cell lines possessed significantly differential expression levels of MMP-2. Quantitative (real‑time) PCR, RT-PCR, and western blot experiments were consistent with such results. The fluorescence detection limit of the probe for MMP-2 was estimated to be 0.05 ng/ml. Therefore, this MMP-2 probe potentially provides a sensitive and specific visual method for cervical cancer screening, diagnosis and prognostic judgment.

摘要

宫颈癌是全球最常见的妇科恶性肿瘤之一。目前,用于宫颈癌筛查的方法存在诸多弊端。基质金属蛋白酶-2(MMP-2)在正常宫颈上皮细胞中含量低或不存在,而在宫颈上皮内瘤变和宫颈癌中过表达,这为基于MMP-2检测宫颈癌提供了可能。有望开发出一种对宫颈癌检测具有高灵敏度和特异性的策略,以提高治愈率。通过MMP-2底物肽GPLGVRGKGG将荧光团[7-(二乙氨基)-2-氧代-2H-色烯-3-琥珀酰亚胺酯]与猝灭剂{4-[4-(二甲氨基)苯基偶氮]苯甲酸N-琥珀酰亚胺酯}连接,制备了一种新型的基于香豆素的靶向MMP-2的荧光共振能量转移探针,用于通过细胞成像检测宫颈癌。宫颈癌细胞系呈现绿色荧光,强度顺序为CaSki > SiHa > C33A > HeLa,这表明宫颈癌细胞系中MMP-2的表达水平存在显著差异。定量(实时)PCR、RT-PCR和蛋白质印迹实验与这些结果一致。该探针对MMP-2的荧光检测限估计为0.05 ng/ml。因此,这种MMP-2探针可能为宫颈癌筛查、诊断和预后判断提供一种灵敏且特异的可视化方法。

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